PROJECT SUMMARY The mechanism(s) by which infection-mediated inflammatory processes drive progressive lung disease remains elusive. Novel preliminary data supports an evolutionary selective metabolism pathway that may explain how the lung defends against pathogen colonization without harming itself in the process. We have discovered that mammalian selenocysteine containing thioredoxin reductase (Sec-TrxR), an enzyme found in the lung, can detoxify the haloperoxidase produced hypothiocyanate (HOSCN) and recycle it back to -SCN. We hypothesize that cysteine-containing thioredoxin reductase (Cys-TrxR) from prokaryotic bacteria such as E. coli and P. aeruginosa cannot detoxify HOSCN due to differences in enzyme structure, especially the lack of a C-terminal selenocysteine residue found only in higher eukaryotes. Cystic fibrosis (CF) is due to mutations in the cystic fibrosis transmembrane conductance regulator (cftr) gene that encodes for an apical membrane anion transporter protein that transports a number of anion species including -SCN. CF subjects have chronic lung infections that contribute to morbidity and mortality associated with this genetic disease. The CFTR protein transports -SCN into the airway surface fluid and we hypothesize that this is a critical feature of the aberrant host defense associated with CF lung disease. -SCN can directly react with HOCl producing HOSCN which can be metabolized by the host but not the pathogen. We provide preliminary data supporting the ability of mammalian Sec-TrxR to detoxify HOSCN and the inability of prokaryotic Cys-TrxR to detoxify HOSCN. We have published in vitro and in vivo data on the importance of CFTR in maintaining airway fluid -SCN levels and preliminary studies that inhaled -SCN dramatically improves morbidity and decreases lung inflammation in a P. aeruginosa lung infection mouse models. Novel data is presented that a SCN analog, selenocyanate (?SeCN), is more potent antimicrobial than SCN, but shares all the selective detoxification benefits of -SCN. The working hypothesis is that in CF the lung cannot use -SCN optimally resulting in haloperoxidases make more hypochlorite (HOCl) which is not selectively detoxified and produces more lung damage and inflammation. Specific aim 1 will determine the chemical mechanism by which Sec-containing TrxR resists inactivation by HOSCN/HOSeCN oxidation and Cys-containing TrxR are inactivated by HOSCN/HOSeCN. Specific aim 2 will examine CFTR?s and Sec-TrxR?s role in the importance of ?SCN/HOSCN -mediated lung host defense against bacterial infection. Specific aim 3 Examine -SeCN as a potential therapy for treatment of lung inflammation and infection using wild type and CFTR KO and ?-ENaC Tg mouse models. The major innovations in this proposal are: 1) our novel discovery of selective detoxification of HOSCN/HOSeCN by mammalian Sec-TrxR that allows the lung to defend against pathogens without harming self; 2) novel targeting of prokaryotic Cys-TrxR by HOSCN/HOSeCN; 3) ?SCN/-SeCN may improve neutrophil and macrophage host defense functions; 4) improvement in lung infection models with nebulized -SeCN; and 5) proof of principle studies to bring -SeCN therapy into the clinic.